Impact of ribavirin on the genetic diversity of the virus of Hepatitis C

A ribavirina é um agente antiviral de amplo espectro com inúmeras aplicações clínicas contra patógenos virais, que pode causar aumento nas taxas de mutação e, potencialmente, resultar na extinção da população viral em um processo denominado mutagênese letal. Diversas abordagens têm sido propostas para compreender esse fenômeno. Foi aprovada pela primeira vez em 1980 para uso em seres humanos como tratamento para infecção grave pelo vírus sincicial respiratório em crianças e tem sido um componente integrante da terapia da infecção pelo HCV desde 2001. Com o objetivo de investigar o impacto da ribavirina na diversidade genética do vírus da hepatite C em pacientes portadores de hepatite C crônica, usamos o sequenciamento NGS para determinar a diversidade genética viral no hospedeiro através da região NS5B do HCV de 7 pacientes tratados com monoterapia de ribavirina em comparação com 11 pacientes sem tratamento. Foram desenhadas três condições diferentes para a etapa de coleta. A primeira, antes de o paciente iniciar o tratamento, a segunda durante a monoterapia de ribavirina e a terceira durante a coadministração com interferon. A avaliação da diversidade genética viral intra-hospedeiro é essencial para compreender a dinâmica evolutiva dos vírus que tem implicações importantes para a persistência, patogênese, respostas imunológicas, transmissão e o desenvolvimento de vacinas e estratégias antivirais bem sucedidas. Este estudo observou dois padrões na diversidade apresentada pelos pacientes que fizeram monoterapia de ribavirina, o primeiro representado por um declínio, enquanto o segundo está mais próximo de um equilíbrio na população viral. O grupo sem tratamento apresentou três perfis diferentes, um declínio, um equilíbrio e um aumento ao longo dos 30 dias. Concluímos que o impacto causado pela ribavirina ao tratamento não é necessariamente, um aumento da diversidade, ou seja, não foi observado um efeito mutagênico viral nos pacientes infectados com o genótipo 1a e 1b. A diversidade ou a ausência dela não influenciou na resposta ao tratamento. A população viral no hospedeiro pode sofrer influências referentes à pressão seletiva tanto do tratamento quanto da imunidade do hospedeiro.Ribavirin is a broad-spectrum antiviral agent with numerous clinical applications against viral pathogens. The drug can increase mutation rates and lead to extinction of the viral population in a process called lethal mutagenesis. Several approaches have been proposed to understand this phenomenon. It was first approved for medical purposes in 1980, as a treatment for severe respiratory syncytial virus infection in children. Since 2001, it has been a component of the therapy for HCV infection. Aiming to investigate the impact of ribavirin on genetic diversity of hepatitis C virus in patients with chronic hepatitis C. NGS sequencing was employed to determine the viral genetic diversity in the host, using the NS5B region of the HCV from 7 patients treated with ribavirin monotherapy, in comparison with 11 untreated patients. Three different conditions were designed for the sample collection stage. The first, before the patient starts treatment, the second during ribavirin monotherapy and the last during co-administration with interferon. The assessment of intra-host viral genetic diversity is essential to understand the evolutionary dynamics of important viruses, implicating in persistence, pathogenesis, immune responses, transmission and the development of successful vaccines and antiviral strategies. This study investigated two patterns in the viral diversity exhibited by patients taking ribavirin monotherapy. The first was represented by a population decline, while the second is closer to equilibrium in the viral population. The untreated group had three different profiles: a decline, equilibrium and an increase over 30 days. We conclude that the impact caused by ribavirin in the treatment is not necessarily an increase in diversity, i. e., a viral mutagenic effect was not observed in patients infected with genotype 1a and 1b. The diversity or its absence did not influence the response to treatment. The viral population in the host may be influenced by the selective pressure of both the treatment and the host's immunity.Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)258/201

Viral Metagenomics for the Identification of Emerging Infections in Clinical Samples with Inconclusive Dengue, Zika, and Chikungunya Viral Amplification

Viral metagenomics is increasingly being used for the identification of emerging and re-emerging viral pathogens in clinical samples with unknown etiology. The objective of this study was to shield light on the metavirome composition in clinical samples obtained from patients with clinical history compatible with an arboviral infection, but that presented inconclusive results when tested using RT-qPCR. The inconclusive amplification results might be an indication of the presence of an emerging arboviral agent that is inefficiently amplified by conventional PCR techniques. A total of eight serum samples with inconclusive amplification results for the routinely tested arboviruses—dengue (DENV), Zika (ZIKV), and Chikungunya (CHIKV) obtained during DENV and CHIKV outbreaks registered in the state of Alagoas, Northeast Brazil between July and August 2021—were submitted to metagenomic next-generation sequencing assay using NextSeq 2000 and bioinformatic pipeline for viral discovery. The performed bioinformatic analysis revealed the presence of two arboviruses: DENV type 2 (DENV-2) and CHIKV with a high genome coverage. Further, the metavirome of those samples revealed the presence of multiple commensal viruses apparently without clinical significance. The phylogenetic analysis demonstrated that the DENV-2 genome belonged to the Asian/American genotype and clustered with other Brazilian strains. The identified CHIKV genome was taxonomically assigned as ECSA genotype, which is circulating in Brazil. Together, our results reinforce the utility of metagenomics as a valuable tool for viral identification in samples with inconclusive arboviral amplification. Viral metagenomics is one of the most potent methods for the identification of emerging arboviruses

Metagenomic insights into the plasma virome of Brazilian patients with prostate cancer

Growing evidence suggests that metavirome changes could be associated increased risk for malignant cell transformation. Considering Viruses have been proposed as factors for prostate cancer induction. The objective of this study was to examine the composition of the plasma metavirome of patients with prostate cancer. Blood samples were obtained from 49 male patients with primary prostate adenocarcinoma. Thirty blood donors were included as a control group. The obtained next-generation sequencing data were analyzed using a bioinformatic pipeline for virus metagenomics. Viral reads with higher abundance were assembled in contigs and analyzed taxonomically. Viral agents of interest were also confirmed by qPCR. Anelloviruses and the Human Pegivirus-1 (HPgV-1) were the most abundant component of plasma metavirome. Clinically important viruses like hepatitis C virus (HCV), cytomegalovirus and human adenovirus type C were also identified. In comparison, the blood donor virome was exclusively composed of torque teno virus types (TTV) types. The performed HPgV-1 and HCV phylogeny revealed that these viruses belong to commonly detected in Brazil genotypes. Our study sheds light on the plasma viral abundance in patients with prostatic cancer. The obtained viral diversity allowed us to separate the patients and controls, probably suggesting that malignant processes may influence virome composition. More complex and multiple approach investigations are necessary to examine the likely causal relationship between metavirome and its nvolvement in prostate cancer

Nucleocapsid (N) Gene Mutations of SARS-CoV-2 Can Affect Real-Time RT-PCR Diagnostic and Impact False-Negative Results

The current COVID-19 pandemic demands massive testing by Real-time RT-PCR (Reverse Transcription Polymerase Chain Reaction), which is considered the gold standard diagnostic test for the detection of the SARS-CoV-2 virus. However, the virus continues to evolve with mutations that lead to phenotypic alterations as higher transmissibility, pathogenicity or vaccine evasion. Another big issue are mutations in the annealing sites of primers and probes of RT-PCR diagnostic kits leading to false-negative results. Therefore, here we identify mutations in the N (Nucleocapsid) gene that affects the use of the GeneFinder COVID-19 Plus RealAmp Kit. We sequenced SARS-CoV-2 genomes from 17 positive samples with no N gene detection but with RDRP (RNA-dependent RNA polymerase) and E (Envelope) genes detection, and observed a set of three different mutations affecting the N detection: a deletion of 18 nucleotides (Del28877-28894), a substitution of GGG to AAC (28881-28883) and a frameshift mutation caused by deletion (Del28877-28878). The last one cause a deletion of six AAs (amino acids) located in the central intrinsic disorder region at protein level. We also found this mutation in 99 of the 14,346 sequenced samples by the Sao Paulo state Network for Pandemic Alert of Emerging SARS-CoV-2 variants, demonstrating the circulation of the mutation in Sao Paulo, Brazil. Continuous monitoring and characterization of mutations affecting the annealing sites of primers and probes by genomic surveillance programs are necessary to maintain the effectiveness of the diagnosis of COVID-19

Correction: Lesbon et al. Nucleocapsid (N) Gene Mutations of SARS-CoV-2 Can Affect Real-Time RT-PCR Diagnostic and Impact False-Negative Results. <i>Viruses</i> 2021, <i>13</i>, 2474

The authors hereby request the inclusion of two authors (Olivia Teixeira and Maria Cristina Nonato) in the recently published article in Viruses entitled “Nucleocapsid (N) gene mutations of SARS-CoV-2 can affect real-time RT-PCR diagnostic and impact false-negative results” [...